A stem cell is normally taken to mean an undifferentiated cell capable of differentiating into all types of mature functional cells constituting a body. For example, a hematopoietic stem cell can differentiate into various corpuscular cells. An embryonic stem (ES) cell derived from an embryo has pluripotency to differentiate and develop into all types of organs, tissues and cells that form a body.
A mouse ES cell line constructed in 1981 has provided a technique and paradigm for developing a human ES cell. The development of the ES cell has been studied using a mouse teratocarcinoma, a tumor that occurs in a gonad of a closely bred mouse strain (Evans & Kaufman, Nature, 292:154-156 (1981)).
Bongso et al. reported a method for culturing and maintaining cells isolated from a human embryo derived from in vitro fertilization for a short-term period (Bongso et al., Human Reproduction, 9:2110-2117 (1994)). The cells isolated by Bongso et al. had a morphology expected in a pluripotent stem cell; however, they could not be cultured for a long-term period apparently because a proper feeder layer was not used.
Primate ES cells have been prepared from a blastocyst of a rhesus monkey and a marmoset monkey. The primate ES cells are diploid, and very similar to a human ES cell.
The study of ES cells prepared from a monkey and a human has suggested that a pluripotent stem cell might be derived from a human blastocyst, although the ES cells from the monkey and the human are somewhat different from that of a mouse in terms of phenotype (Thomson et al., Proc. Natl. Acad. Sci. USA, 92:7844-7848 (1995)).
The characteristic features of human pluripotent ES cells developed by Thomson et al. in 1998 (Thomson et al., Science, 282:1145-1147 (1998)) are as follows:
(1) expression of stage-specific embryonic antigen-3 (SSEA-3), stage-specific embryonic antigen-4 (SSEA-4), tumor rejection antigen 1-60 (TRA-1-60), tumor rejection antigen 1-81 (TRA-1-81), and alkaline phosphatase;
(2) high telomerase activity;
(3) differentiation into three types of blastodermal cells when injected into mice;
(4) dependency on feeder cells; and
(5) no response to a human leukemia inhibitory factor (hLIF).
Thomson et al. obtained the above ES cells from a blastocyst donated by a couple under sterility treatment. Specifically, a trophectoderm known to inhibit the establishment of an ES cell was removed immunosurgically, an inner cell mass (ICM) was plated on a fibroblast feeder layer derived from a mouse embryo, and the ICM was replated on another feeder layer after a short attachment and expansion period. Thomson's method was not significantly different from the mouse ES cell protocol in terms of the medium or culture system; and yet a relatively high success rate was achieved.
The isolation of human pluripotent ES cells and breakthroughs in somatic cell nuclear transfer (SCNT) in mammals (Solter, Nat. Rev. Genet., 1:199-207 (2000)) have raised the possibility of performing human SCNT to generate virtually unlimited sources of undifferentiated cells for research, with potential applications in tissue repair and transplantation medicine. This concept, known as “therapeutic cloning,” employs a nuclear transfer of a somatic cell into an enucleated oocyte (Lanza et al., Nat. Med., 5:975-977 (1999)). Previous studies on such therapeutic cloning dealt with the production of bovine ES-like cells (Cibelli et al., Nat. Biotechnol., 16:642-646 (1998)) and mouse ES cells from ICMs of cloned blastocysts (Munsie et al., Curr. Biol., 10:989-992 (2000); Wakayama et al., Science, 292:740-743 (2001)) and development of cloned human embryos until 8 to 10 cell stages (Cibelli et al., J. Regen. Med., 2:25-31 (2001)).
Although several reports have indicated that an ES cell line can be established by employing a non-human mammalian oocyte, no ES cell line developed from a human oocyte utilizing the nuclear transfer technology has been reported yet.